Several publications are referenced in this application in parentheses in order to more fully describe the state of the art to which this invention pertains. The disclosure of each of these publications is incorporated by reference herein.
P-glycoprotein, the product of the MDR1 gene, was the first ABC transporter shown to confer resistance to cytotoxic agents. Pgp functions as an ATP-dependent efflux pump that reduces the intracellular concentration of a variety of chemotherapeutic agents by transporting them across the plasma membrane (1). The multidrug resistance phenotype associated with overexpression of Pgp is of considerable clinical interest because natural product drugs are second only to alkylating agents in clinical utility, and many effective chemotherapeutic regimens contain more than one natural product agent. More recently, we and others have reported transfection studies indicating that MRP, another ABC family transporter, confers a multidrug resistance phenotype that includes many natural product drugs, but is distinct from the resistance phenotype associated with Pgp (2-6). MRP shares only limited amino acid identity with Pgp, and this is reflected in the different substrate specificities of the two transporters. In contrast to Pgp, MRP can transport a wide range of anionic organic conjugates, including glutathione S-conjugates (7). In addition to Pgp and MRP there may be other transporters that are involved in cytotoxic drug resistance. In the case of natural product drugs, resistant cell lines have been described that display a multidrug resistant phenotype associated with a drug accumulation deficit, but do not overexpress Pgp or MRP (8). ABC transporters have also been linked to cisplatin resistance, and several lines of evidence suggest the possibility that pumps specific for organic anions may be involved: 1) decreased cisplatin accumulation is consistently observed in cisplatin resistant cell lines (9); 2) cisplatin is conjugated to glutathione in the cell, and this anionic conjugate is toxic in an in vitro biochemical assay (10); and 3) biochemical studies using membrane vesicle preparations have shown that cisplatin resistant cells lines have enhanced expression of an ATP-dependent transporter of CDDP-glutathione and other glutathione S-conjugates such as the cystinyl leukotriene LTC4 (11, 12). These data thus suggest that an organic anion transporter may contribute to cisplatin resistance by exporting CDDP-glutathione. While MRP is an organic anion transporter, the reported drug resistance profile of MRP-transfected cells does not extend to this agent (5, 6), and to date only one cisplatin resistant cell line has been reported to overexpress MRP (13). This suggests that organic anion transporters other than MRP may contribute to cisplatin resistance. Consistent with this possibility, the canalicular multispecific organic anion transporter, cMOAT, an MRP-related transporter that functions as the major organic anion transporter in liver, has been reported to be overexpressed in cisplatin resistant cell lines (14, 15). A more direct link between cMOAT and cytotoxic drug resistance is suggested by a recent report in which transfection of a cMOAT antisense construct into a liver cancer cell line resulted in sensitization to cisplatin, daunorubicin and other cytotoxic agents (16).
Clearly, a need exists for identifying the essential components and mechanisms giving rise to drug resistance and the transport of anticancer agents out of the tumor cell. The elucidation of these mechanisms may be used to advantage for the design of efficacious chemotherapeutic agents.